L-Arginine (Arg) is a conditionally essential amino acid, naturally found in dietary protein. It is converted to nitric oxide (NO) (Palmer et al. Nat Med 1987; 327:524-526; Moncada et al. N Engl J Med 1993; 329:2002-2012; Kam et al. Anaesthesia 1994; 49:515-521) and bronchodilator (Zoritch et al. Arch Dis Child 1995; 72:259-262; Gaston et al. Am J Respir Crit Care Med 1994; 149:538-551), a potent vasodilator, by a family of enzymes known as nitric oxide synthase (NOS). NO is an essential molecule that plays a role in a broad range of functions from vascular regulation, neurotransmission (Moncada et al. 1993, supra), host defense, and cytotoxicity (Nathan et al. Proc Natl Acad Sci 2000; 97:8841-8848) to physiologic control of airways (Gaston et al. 1994, supra). Under conditions of low L-arginine concentration, nitric oxide synthase is uncoupled and reduces oxygen (O2) to superoxide (O2−) instead of generating nitric oxide (Xia et al. Proc Natl Acad Sci 1996; 93:6770-6774; Dias-Da-Motta et al. Brit J Haematol 1996; 93:333-340). Nitric oxide reacts rapidly with superoxide to form reactive nitric oxide species (RNOS) that could lead to worsening inflammation, oxidative stress and cellular damage (Demiryurek et al. Pharm Toxicology 1998; 82:113-117).
Recently, expression of inducible NO synthase, the enzyme that catalyzes the production of NO from L-Arg, has been found in the epithelium of asthmatic patients but not in healthy non-asthmatic patients (Hamid et al. Lancet 1993; 342:1510-1513: Nijkamp et al. Arch Int Pharmoocodyn 1995; 329:81-96). Asthmatics have exhaled air NO levels that are 3.5 times higher than non-asthmatics, which are correlated with decrease in FEV1 and are affected by therapy Kharitonov et al. Eur Respir J 1995; 8:295-7). Blocking of NO production by L-Arg analogues results in an increase in allergen-induced bronchoconstriction (Ricciardolo et al. Lancet 1996; 348:374-377). A deficiency of NO is involved in airway hyperreactivity (Meurs et al. Br J Pharmacol 1999; 126:559-562). Although asthma is clearly a multifactorial disease, there is some evidence that NO may play an important role in disease pathogenesis (Sanders et al. Am J Respir Cell Mol Biol 1999; 21:147-149). For reviews, see, e.g., Dweik Cleve Clin J Med. 2001 June; 68(6):486, 488, 490, 493; Gianetti et al. Eur J Clin Invest. 2002 August; 32(8):628-35.
Arginase is an enzyme that hydrolyzes Arg to produce ornithine and urea, (Boucher et al. Cell Mol Life Sci 1999; 55:1015-1028) however, in the presence of nitric oxide synthase (NOS), arginine is converted to nitric oxide (NO) and citrulline (Moncada et al. 1993, supra). The expression of arginase can be induced by a variety of cytokines involved in the inflammatory process (Solomons et al. Pediatr 1972; 49:933), particularly the Th2 cytokines. (Mori et al. 2000. Relationship between arginase activity and nitric oxide production. In L. Ignarro, editor. Nitric Oxide. Biology and Pathology. Academic Press, San Diego. 199-208.).
Increased serum arginase activities have been reported in patients with SCD at steady-state (Waugh et al. Nutritional Research 1999; 19:501-518.), as well as in an asthma animal model (Meurs et al. Br J Pharmacol 2002; 136:391-398). Arginase activity is elevated in SCD patients with pulmonary disease (Morris et al. Am J Respir Crit Care Med 2003; 168:63-69; Morris et al. 2002. Elevated serum arginase activity in patients with sickle cell disease and pulmonary hypertension. The 30th Anniversary of the National Sickle Cell Program, Washington, D.C.). Plasma arginase activity appears to be related to hemolysis, associated with several markers of hemolytic severity, including LDH (r=0.44, p<0.001), AST (r=0.39, p<0.002), reticulocyte count (r=0.25, p<0.001), and Hct (r=−0.25, p<0.001) (Morris et al, Erythrocyte arginase release during hemolysis contributes to endothelial dysfunction and pulmonary hypertension, 27th Annual Meeting of the National Sickle Cell Disease Program, Los Angeles, Calif.; April 2004).
Arginase controls the metabolism of arginine into ornithine, which in turn gives rise to proline and polyamines (Mori et al. 2000, supra; Morris Annu Rev Nutr 2002; 22:87-105; Morris 2000. Regulation of arginine availability and its impact on NO synthesis. Nitric Oxide. Biology and Pathobiology. Academic Press, San Diego. 187-197; Mori et al. Biochem Biophys Res Commun 2000; 275:715-719). These downstream products of arginase activity may play a significant role in the pathogenesis of asthma, pulmonary hypertension and other inflammatory conditions, since proline is involved in collagen formation (Kershenobich et al. J Clin Invest 1970; 49:2246-2249; Albina et al. J Surg Res 1993; 55:97-102) and lung fibrosis (Endo et al. Am J Physiol Lung Cell Mol Physiol 2003; 285:L313-L321), processes that occur in airway wall thickening and airway remodeling (Tanaka et al. Inflamm Res 2001; 50:616-624: Elias et al. J Clin Invest 1999; 104:1001-1006; Elias et al. J Clin Invest 2003; 111:291-297; Busse et al. N Engl J Med 2001; 344:350-362).
Arginine, a safe dietary supplement, has already demonstrated potential for therapeutic utility in several disease processes. (Pieper Hypertension 1998; 31:1047-1060; Lerman et al. Circulation 1998; 97:2123-2128 Perrine et al. N Engl J Med 1993; 328:81-6; Maxwell et al. Current Opinion in Nephrology and Hypertension. 133; Creager et al. J Clin Invest 1992; 90:1248-53; Drexler et al. Lancet 1991; 338:1546-50). In animal studies, inhalation of low doses of L-Arg has completely blocked hyperresponsiveness of reactive airways (Nijkamp et al. 1995, supra; Folkerts et al. J Clin Invest 1995; 94:26-30), and inhaled L-Arg also improves pulmonary functions of cystic fibrosis patients (CF) (Solomons et al. Pediatr 1971; 47:384-390; Solomons et al. Pediatr 1972; 49:933). When tested in a mouse model of allergic asthma, oral administration of L-Arg was reported to aggravate allergen-induced eosinophilic airway inflammation (Takano et al. J Pharmacol Exp Ther 1998 August; 286(2):767-71).
Use of L-Arg is suggested for treatment of cystic fibrosis (Busch-Petersen et al. Z Erkr Atmungsorgane 143:140-7 (1975)); treatment of exercise induced pulmonary hemorrhage in horses (U.S. Pat. No. 6,027,713); and treatment of pulmonary hypertension (U.S. Pat. Nos. 5,217,997; 6,127,421; Nagaya et al. Am J Respir Crit Care Med 163:887-81 (2001); Cheng et al. Hua Xi Yi Ke Da Xue Xue Bao 27:68-70 (1996)).
Use of NO to treat asthma is discussed in Nakagawa et al. J Pediatr. 2000 July; 137(1):119-22; and Rossaint et al. Eur Heart J 1993 November; 14 Suppl I:133-40). The arginase inhibitor N-hydroxy-L-arginine (NOHA) has been tested in a model of asthma (see, e.g., Meurs et al., Br J Pharmacol June 2002, 136(3):391-8, describing administration of an arginase inhibitor in a guinea pig model of allergic asthma; and Meurs et al. Br J Pharmacol 130:1793-8 (2000, describing arginase inhibitors in a perfused guinea pig trachea model)). Use of NO to treat pneumonia has been discussed (see, e.g., Kimura et al. Pediatr Int 2002 August; 44(4):451-2; Ho et al. J R Soc Med 2002 January; 95(1):35-7; Bugge et al. Eur J Anaesthesiol 2000 April; 17(4):269-72; Hoehn et al. Respiration 1998; 65(6):477-80; Blomqvist et al. Acta Anaesthesiol Scand 1993 January; 37(1):110-4; Jean et al. Crit Care Med 2002 February; 30(2):442-7 and Kannan et al. Indian J Pediatr 1998 May-June; 65(3):333-45).
Although early investigators warned of the deleterious impact of nitric oxide in sickle cell disease (SCD) (Knight et al. Pediatr Pulmonol 1999; 28:205-216), more recent studies support its protective function (Gladwin et al. Semin Hematol 2001; 38:333-342). Similar to asthmatic patients (Lopez da Mata et al. 1998. How does nitrates in blood correlated to exhaled levels in asthma? European Respiratory Conference, Geneva, Switzerland.), SCD patients also have elevated NOx levels at baseline (Rees et al. Br J Haematol 1995; 91:834-7). Serum L-Arg and NOx levels fall, however, during the vaso-occlusive complications of SCD, (Morris et al. J Pediatr Hematol Oncol 2000; 22:515-520) with lowest levels found during acute chest syndrome (pneumonia). Most SCD patients with pulmonary disease have a component of reactive airways that respond to bronchodilators, even though they often do not demonstrate the classical wheezing on physical exam that is usually associated with asthma. Asthma in SCD is often unrecognized and undertreated, and occurs in 30-60% of patients (Minter et al. Am J Respir Crit Care Med 2001; 164:2016-2019). Clinical trials of arginine therapy are now underway for SCD (Morris et al. Brit J Haematol 2000; 111:498-500; Morris et al. 2003, supra).
Magnesium, which can be a dietary supplement, has been described as an adjuvant in combination therapy of asthma with salbumatol (Hughes et al, Lancet 2003; 361:2114-7) or as an asthma intravenous monotherapy (Gurkan et al, Eur J Emerg Med 1999; 6:201-5). Magnesium has also been suggested in infusion therapy of neonatal pulmonary hypertension (Patole et al. Magnes Res 1995; 8:373-88). The effects of oral magnesium in an animal model of pre-eclampsia has been reported (Pandhi et al, Indian J Exp Biol 2002; 40:349-51) and other disease processes that involve endothelial dysfunction (Volpe et al, Scand Cardiovasc J 2003; 37:288-96). Magnesium-induced vasodilation has been reported in animal models of other conditions that involve endothelial-derived nitric oxide (Teragawa et al, Magnes Res 2002; 15:241-6, describing the effects of magnesium in an in vitro canine coronary artery model of endothelial dysfunction). Combined therapy of magnesium and inhaled nitric oxide has shown some promise in an animal model of pulmonary hypertension (Haas et al, Pediatr Int 2002; 44:670-4).
Despite the advances in the field with respect to therapies for conditions such as asthma and sickle cell disease, new therapies are of considerable interest and importance. Furthermore, diagnosis and therapies based upon a more insightful understanding of the underlying mechanisms of these diseases is needed so as to provide a more rationale approach to therapy.
There is a need in the field for improved or alternative therapies for treatment of conditions such as asthma. The present invention addresses these needs.